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Background

B-cell targeted therapies are important strategies in human systemic lupus erythematosus
(SLE). Previous studies have shown that B-cell complement receptor type 2 (CR2/CD21),
along with its C3 activation fragment antigen-bound ligand designated C3d, play essential
roles in the innate-adaptive immune interface and development of antibodies to foreign
antigens. CR2 acts with CD19 to greatly amplify B-cell receptor signals. We hypothesize
that a similar role is played by this receptor-ligand pair in the development of high-affinity
IgG autoantibodies in patients with SLE. Prior gene-targeting studies have suggested,
however, that CR2 expression may be needed to maintain tolerance. These studies are
confounded, however, because not only is CR2 absent but due to murine-specific gene
structures another receptor designated CR1 is also deleted at the same time. CR1 is
a receptor for complement fragment C4b, whose deficiency in humans and murine models
leads to lupus. In addition, a recent report of the first identified human CR2-deficient
individual revealed a humoral immunodeficiency and not an autoimmune phenotype. Other
recent studies have shown in MRL/lpr and (NZB×NZW)F1 mice that the use of soluble CR2 as a potential dominant negative
inhibitor led to a substantial decrease in autoantibody titers.

Methods

To address our hypothesis, we developed novel mAbs that disrupt the CR2-C3d interface
alone, without affecting the interactions of CR1 with C4b. We immunized C3-/- mice with recombinant human C3d, and Cr2-/- mice with recombinant murine CR2.

Results

The resultant human C3d-reactive mAbs inhibited C3d-CR2 binding, did not recognize
intact C3/C3b, and cross-reacted with mouse C3d. Two anti-C3d mAbs, 3d29 and 3d8b,
along with control mAb were pre-injected into mice before sheep red blood cell (SRBC)
immunization. IgG1 responses to SRBC antigen were substantially decreased, consistent
with the interruption in vivo of C3d binding to CR2. One resulting anti-CR2 mAb (4B2, IgG1), which directly blocks binding of C3d to CR2, was injected in wild-type mice and
demonstrated no B-cell depletion but maintenance of blockade of CR2 on the B-cell
surface for at least 1 month. SRBC immunization of mice pre-injected with mAb 4B2
revealed reduced anti-SRBC levels to levels found in immunized Cr2-/- mice. No anti-idiotype antibodies were detected.

Conclusion

We have developed unique tools to characterize in mouse models of human lupus the
pathogenic roles of both the C3d ligand and CR2 components of the CR2-C3d interaction
pair.